Ca2+对角质形成细胞增殖及分化调控作用探究.docVIP

Ca2+对角质形成细胞增殖及分化调控作用探究[摘要]目的：探讨胞外Ca2+对人角质形成细胞(HKC)的增殖与分化的调控作用，为组织工程皮肤的研制提供依据。方法：HKC体外培养，应用形态学、免疫细胞化学、MTT等方法检测胞外Ca2+对HKC增殖与分化的影响，并利用钙通道阻滞剂SKF 96365来研究其作用的可能途径。结果：Ca2+为0.5mmol/L时，HKC呈单层生长，免疫细胞化学显示HKC呈低分化状态，当Ca2+为1mmol/L与1.5mmol/L时，细胞在3天后出现复层生长，免疫细胞化学显示细胞K19角蛋白表达减少，Ca2+为2.0mmol/L时表现出一定的细胞毒性作用，SKF 96365可逆转Ca2+的促增殖与分化作用。结论：胞外Ca2+在一定范围内具有促HKC分化与增殖的作用，并且该作用可以被SKF 96365所阻滞。 [关键词]Ca2+；角质形成细胞；组织工程；钙通道阻滞剂 [中图分类号]Q813.1[文献标识码]A[文章编号]1008-6455（2007）09-1181-03 The regulative effect of Ca2+ on proliferation and differentiation of epidermal keratinocytes SUN Qiang,CHAI Jia-ke,LIANG Li-ming,YANG Hong-ming,DU Jia-mei,WANG Li-huan (Department of Plastic and Burns Surgery,First Affiliated Hospital,Chinese PLA General Hospital,Beijing 100037,China) Abstract:ObjectiveTo investigate the regulative effect of Ca2+ on growth and differentiation of keratinocytes which is the base of tissue engineering skin.MethodsKeratinocytes were cultured with H-KGM contained different levels of Ca2+ or Ca2+ entry blocker. The result were detected by morphology , immunocytochemistry and MTT assay for two weeks.ResultsThe cell were in monolayer in H-KGM at level of 0.5mmol/L Ca2+，However, as concentration of Ca2+ was 1mmol/L or 1.5mmol/L, cell grown up rapidly and became stratum after 1-3d of culture, the expression of keratin 19 decreased. We also found the cytotoxicity of 2.0mmol/L Ca2+, this effect was markly suppressedby Ca2+ entry blocker.ConclusionCa2+ can promote the proliferation and differentiation of kerationcyte, and this effect can be depressed by the Ca2+ entry blocker. Key words: Ca2+ Keratinocyte;tissue engineer;Ca2+ entry blocker 随着组织工程研究工作的不断深入，更多的学者[1]认识到目前组织工程研究工作的重点是如何解决种子细胞的调控问题。HKC是构建组织工程皮肤重要的种子细胞，研究表明，HKC的增殖与分化是受多因素调节的复杂过程。Ca2+作为体内活跃的阳离子，参与了机体多种生理性与病理性活动，有资料[2]显示，Ca2+具有促表皮细胞增殖与分化的作用，但Ca2+对HKC的调节作用与其剂量之间具体的关系，以及钙通道阻滞剂对这种调节作用的影响仍存在众多争议。本文以体外培养的HKC为研究对象，观察Ca2+对其增殖与分化的影响，旨在为组织工程皮肤的研制与应用奠定一定的理论基础。 1材料与方法 1.1 材料 1.1.1 实验对象：HKC取自包皮环切术与烧伤整形植皮手术剩余皮片，患者知情同意。 1